The overall objective of this project is to study mutagenesis in mammalian cells in vitro and to demonstrate how such systems can be useful in predicting the nature of putative genetic alterations inflicted by environmental agents on the human community. The system we are developing differs from most mammalian cell mutagenesis studies in two fundamental respects. Instead of studying the genetic effects of agents at a single locus, we look for variants for over 50 different gene products within a single cell. Secondly, our system can distinguish between genetic changes which modify the gene product (structural gene mutations) and those which alter the expression of a locus (control or regulational events). Chinese hamster ovary (CHO) cells are used. Cells are exposed to mutagen (UV or MNNG in these initial experiments), allowed to go through two divisions to segregate out mutant strands and then cloned randomly. Each clone is grown to 60 x 10 to the 6th power cells. A sample is kept in culture while the remainder is homogenized. Supernatants are then subjected to electrophoresis and gels are histochemically stained for the products of over 50 enzyme loci. The following alterations in the banding patterns of the loci products are sought--electrophoretic shift consistent with the subunit structure of the enzyme implying an alteration in the structural gene coding for the protein, presence of a gene product not normally expressed in the cell line, or loss of expression of a gene product normally produced. Any clone shown to be variant is then subcloned to determine the heritability of any variants. BIBLIOGRAPHIC REFERENCES: Siciliano, M J. and R. M. Humphrey, 1976a. Stable, inherited modifications of gene expression in mammalian somatic cells after exposures to mutagen. First Int. Cong. Cell Biology, in press (Abstract). Siciliano, M. J. and R. M. Humphrey, 1976b. Functional genetic diploidy in CHO cells as demonstrated by induced electrophoretic enzyme variants. Genetics, in press. (Abstract)